Characterization of a cellobiose dehydrogenase from Humicola insolens.
The major cellobiose dehydrogenase (oxidase) (CBDH) secreted by the soft-rot thermophilic fungus Humicola insolens during growth on cellulose has been isolated and purified. It was shown to be a haemoflavoprotein with a molecular weight of 92 kDa and a pI of 4.0, capable of oxidizing the anomeric carbon of cellobiose, soluble cellooligosaccharides, lactose, xylobiose and maltose. Possible electron acceptors are 2,6-dichlorophenol-indophenol (DCPIP), Methylene Blue, 3,5-di-t-butyl-1,2-benzoquinone, potassium ferricyanide, cytochrome c and molecular oxygen. The oxidation of the prosthetic groups by oxygen was monitored at 449 nm for the flavin group and at 562 nm for the haem group. The curves were very similar to those of the cellobiose dehydrogenase from Phanerochaete chrysosporium, suggesting a similar mechanism. The pH-optima for the oxidation varied remarkably depending on the electron acceptor. For the organic electron acceptors, the pH-optima ranged from pH 4 for Methylene Blue to pH 7 for DCPIP and the benzoquinone. In the case of the FeIII-containing electron acceptors, the enzyme displayed alkaline pH-optima, in contrast to the properties of cellobiose dehydrogenases from Phanerochaete chrysosporium and Myceliophthora (Sporotrichum) thermophila. The enzyme has optimal activity at 65 degrees C.
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Lovens kemiske Fabrik, Ballerup, DK-2750, Denmark.
Christensen
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MH
Schülein
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D000596:Amino Acids / Q000032:analysis; D002237:Carbohydrate Dehydrogenases / Q000302:isolation & purification* / Q000378:metabolism; D002384:Catalysis; D005656:Fungal Proteins / Q000302:isolation & purification / Q000378:metabolism; D006420:Hemeproteins / Q000302:isolation & purification / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D003904:Mitosporic Fungi / Q000201:enzymology*; D010084:Oxidation-Reduction; D013057:Spectrum Analysis; D013696:Temperature
D000596:Amino Acids; D005656:Fungal Proteins; D006420:Hemeproteins; D002237:Carbohydrate Dehydrogenases; C019859:cellobiose-quinone oxidoreductase